Regulation of basal and tumour necrosis factor α-induced fibroblast gene expression by peptidylarginine deiminase 2

Fibroblasts exhibit organ-specific function and in response to tumour necrosis factor α (TNFα) can produce a large amount of inflammatory mediators, including interleukin-6 (IL-6), partly via a leukaemia inhibitory factor (LIF) autocrine loop, thereby playing a critical pathogenic role in several organ-specific autoimmune diseases, such as rheumatoid arthritis (RA). Peptidylarginine deiminases (PADs), including PAD1-4 and PAD6 in humans, irreversibly convert peptidylarginine to non-coding peptidylcitrulline, a process also known as citrullination. Although the behaviour of fibroblasts is influenced by the citrullination status of extracellular matrix proteins, whether PADs cell-autonomously regulate gene expression of fibroblasts is still unclear. Interestingly heightened PAD activity is strongly associated with RA, which often co-occurs with interstitial lung disease (ILD). The molecular mechanism mediating the unique lung-synovium connection in RA is still poorly understood. Here we show that PAD2 transcript levels in lung fibroblasts (LFs) increased with age, a trend not observed with other PADs or in fibroblasts from other organs. PAD2 deficiency reduced the expression of Podoplanin and Tumor necrosis factor receptor superfamily member 1B (TNFRSF1b) across fibroblasts from all mouse organs and human LFs. Additionally PAD2 deficiency attenuated the expression of several LIF-dependent and LIF-independent genes, including chemokines and Il6, in LFs after TNFα stimulation. Particularly a reduction in TNFα-induced interleukin-6 (IL-6) expression was also detected in fibroblasts from synovium but not in fibroblasts from other organs. These findings demonstrate that PAD2 regulates TNFα-induced gene expression of fibroblasts depending on their organ origin, suggest a novel pathogenic role for PAD2 in RA and offer a plausible explanation for the observed lung-synovium connection. KEY POINTS: Fibroblasts in response to tumour necrosis factor α (TNFα) produce a high level of inflammatory cytokine interleukin-6 (IL-6) that drives the joint and often lung inflammation seen in rheumatoid arthritis (RA), which is characterized by autoimmunity against citrullinated antigens produced by peptidylarginine deiminases (PADs). Fibroblast-derived PAD2 can citrullinate extracellular matrix proteins (ECMPs), whereas the citrullination status of ECMPs reciprocally influences the behaviour of fibroblasts. We examined the cell-autonomous role of PAD2 in fibroblast gene expression and found that PAD2 levels uniquely increase with age in lung fibroblasts (LFs) but not fibroblasts from other organs. PAD2 maintains the expression of Podoplanin and Tumor necrosis factor receptor superfamily member 1B (TNFRSF1b) in fibroblasts across multiple organs but promotes basal and TNFα-induced IL-6 expression in lung and synovial fibroblasts. These results suggest a novel pathogenic role for fibroblast-derived PAD2 in RA and offer a plausible explanation for the unique tissue tropism of RA.